1. Field of the Invention
The invention relates to a method for increasing the hybridization in cells in vitro and for increasing the production and secretion of substances, formed by such cell hybrids, a method for increasing the production of monoclonal antibodies in vitro by means of a hybridoma technique and endothelial cells and/or supernatant of endothelial cells.
2. Brief Description of the Prior Art
The introduction of hybrids between two different cells appears to be successful in the in vitro synthesis and secretion of various cell products. Furthermore, this technique is largely used in gene mapping. It was shown (Kohler, G. and Milstein, C: Nature, Vol. 256, p. 495,-1975-) to permit the production of monoclonal antibodies (thus pure antibodies of one specificity directed to a single antigenic determinant) rather than a mixture of antibodies thus with several (undesired) specificities, as obtained by conventional immunization in vivo. Futhermore, by means of the hybridization of human cells the production of human monoclonal antibodies may be obtained.
The technique of the production of monoclonal antibodies comprises essentially three steps:
(1) Fusion of myeloma cell line cells with lymphocytes obtained from an animal immunized with the desired antigen in order to obtain hybridomas secreting the desired antibodies and growing permanently in vitro.
(2) Selection of the desired hybridoma clone by means of culturing at the single cell level (limiting dilution).
(3) Keeping alive, splitting and maintaining the function of the selected monoclonal cell line in vitro during a prolonged time.
It appeared that all these steps present a number of problems:
(1) The fusion is usually performed at non constant and relatively low efficiency (number of hybrids obtained is usually too low).
(2) The limiting dilution requires the presence of "feeder" cells; said often provide a strongly varying activity.
(3) The survival of hybridomas obtained by fusion of human cells with mouse-myeloma cells is often very limited in time.
The above-mentioned steps and problems are in common with all the hybrids between different cells.